瞬时转染CD44反义寡核苷酸抑制人胃癌MGC80-3细胞增生并诱导凋亡

Inhibitory and apoptosis-inducing effects of CD44 antisense oligodeoxyribonucleotides on human gastric cancer cell line MGC80-3

目的:研究CD44反义寡核苷酸(CD44ASODN)对人胃癌MGC80-3细胞的增生抑制和诱导凋亡的作用和机制. 方法:设计并合成CD44SODN,脂质体介导转入MGC80-3 胃癌细胞,采用流式细胞术(FACS)检测CD44、Fas的表达及细胞凋亡;RT-PCR法检测CD44mRNA的表达;MTT 法检测细胞增生. 结果:CD44ASODN(1.6μmol/L)明显地抑MGC80-3细胞CD44mRNA和蛋白表达水平.CD44ASODN作用MGC80-3 细胞48 h后,细胞的增生呈现明显的抑制作用,其抑制率为31.0%,72,96 h的抑制率分别为46.3%、49.6% (P<0.01),其增生抑制作用呈时间依赖效应.在CD44配体低分子质量透明质酸存在的环境中,CD44ASODN能显著增高细胞表面Fas分子的表达,表达率从6.7%提高为16.8% (P<0.01),并显著地增加MGC80-3细胞对FasmAb诱导凋亡的敏感性,凋亡率从0增加到26.5%(P<0.01). 结论:CD44反义寡核苷酸通过抑MGC80-3细胞CD44m RNA和蛋白表达,抑制MGC80-3细胞的增生,增高细胞表面Fas的表达及MGC80-3细胞对FasmAb诱导凋亡的敏感性,逆转胃癌细胞的免疫逃逸作用.

AIM: To study the role and mechanism of antisense oligodeoxyribonucleotides (ASODN) on proliferation and apoptosis of MGC80-3 cells. METHODS: Flow cytometry was used to detect CD44, Fas expression and apoptosis of MGC80-3 cells. Reverse transcription polymerase chain reaction (RT-PCR) assay was used to examine CD44 mRNA level; MTT assay was used to detect cell proliferation. RESULTS: CD44 mRNA and CD44 protein expression in MGC80-3 cells was blocked and down-regulated after transfected with CD44ASODN (1.6 μmol/L). CD44ASODN inhibited the growth of MGC80-3 cells with depression ratios of 31.0%, 46.3% (P<0.01), and 49.6% (P<0.01) at 48 h, 72 h, and 96 h respectively in a time-dependant manner. With the existence of hyaluronic acid, CD44ASODN improved Fas expression in MGC80-3 cells from 6.7% to 16.8% (P<0.01). At the same time, it enhanced the susceptibility of MGC80-3 cells to Fas mAb and the apoptotic rate of the cells increased from 0% to 26.5% (P<0.01). CONCLUSION: CD44ASODN can down-regulate the expression of CD44 mRNA and protein, inhibit MGC80-3 cell pro liferation and promote apoptosis induced by Fas mAb.